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The large tegument protein pUL36 is essential for formation of the capsid vertex-specific component at the capsid-tegument interface of herpes simplex virus 1.

Fan WH, Roberts AP, McElwee M, Bhella D, Rixon FJ, Lauder R - J. Virol. (2014)

Bottom Line: In addition, the presence of full-length pUL36 results in weak density that extends the CVSC toward the penton, suggesting either that this extra density is formed directly by pUL36 or that pUL36 stabilizes other components of the vertex-tegument interface.Herpesviruses have complex particles that are formed as a result of a carefully controlled sequence of assembly steps.We show that the largest viral protein, pUL36, which occupies the layer of tegument closest to the capsid, is essential for formation of structurally normal connections to the capsid.

View Article: PubMed Central - PubMed

Affiliation: MRC-University of Glasgow Centre for Virus Research, Glasgow, United Kingdom.

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Protein compositions of HSV-1 WT and mutant virus capsids as determined by Western blot analysis of cytoplasmic C-capsids purified from the cytoplasm of cells infected with WT HSV-1 or the inner tegument deletion mutants ARΔUL36 and FRΔUL37. Serial 2-fold dilutions of each sample were run on a 10% polyacrylamide gel and transferred to Hybond ECL nitrocellulose membranes for analysis. Equal amounts of purified WT virions were loaded for comparative purposes. Proteins were visualized using antibodies MAbE12-E3 (pUL36), M780 (pUL37), MAb203 (pUL17), and MAb166 (pUL25). The major capsid protein pUL19 was monitored as a loading control using antibody DM165. The positions of molecular weight standards (in thousands) are indicated to the right of each panel.
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Figure 2: Protein compositions of HSV-1 WT and mutant virus capsids as determined by Western blot analysis of cytoplasmic C-capsids purified from the cytoplasm of cells infected with WT HSV-1 or the inner tegument deletion mutants ARΔUL36 and FRΔUL37. Serial 2-fold dilutions of each sample were run on a 10% polyacrylamide gel and transferred to Hybond ECL nitrocellulose membranes for analysis. Equal amounts of purified WT virions were loaded for comparative purposes. Proteins were visualized using antibodies MAbE12-E3 (pUL36), M780 (pUL37), MAb203 (pUL17), and MAb166 (pUL25). The major capsid protein pUL19 was monitored as a loading control using antibody DM165. The positions of molecular weight standards (in thousands) are indicated to the right of each panel.

Mentions: pUL17 and pUL25, which make up the CVSC, are essential for packaging and retention of DNA in capsids. Therefore, the greatly reduced size of the CVSCs found on ARΔUL36 capsids was surprising, as both the component proteins would be expected to be present on DNA-containing C-capsids. The small size of the CVSC in these cytoplasmic C-capsids suggests either that the archetypal CVSC seen on WT nuclear C-capsids contains, or is stabilized by, pUL36 or that the UL36 mutant capsids contain altered amounts of pUL17 and pUL25. To investigate the latter possibility, we examined the protein composition of the capsids by Western blotting (Fig. 2). Increasing amounts of each capsid preparation were probed with antibodies against pUL17, pUL19, pUL25, pUL36, and pUL37. As expected, pUL36 and pUL37 were not detected in the ARΔUL36 and FRΔUL37 samples, respectively. ARΔUL36 capsids also lacked pUL37, thereby confirming that complex formation with pUL36 is required for addition of pUL37 to capsids (55). pUL36 and pUL37 were both present in WT capsids, but the amounts were appreciably lower than in WT virions. Importantly, the amounts of pUL17 and pUL25 in ARΔUL36 capsids were similar to those for the WT and FRΔUL37, thereby demonstrating that the reduced size of the CVSC was not caused by loss of either of these proteins.


The large tegument protein pUL36 is essential for formation of the capsid vertex-specific component at the capsid-tegument interface of herpes simplex virus 1.

Fan WH, Roberts AP, McElwee M, Bhella D, Rixon FJ, Lauder R - J. Virol. (2014)

Protein compositions of HSV-1 WT and mutant virus capsids as determined by Western blot analysis of cytoplasmic C-capsids purified from the cytoplasm of cells infected with WT HSV-1 or the inner tegument deletion mutants ARΔUL36 and FRΔUL37. Serial 2-fold dilutions of each sample were run on a 10% polyacrylamide gel and transferred to Hybond ECL nitrocellulose membranes for analysis. Equal amounts of purified WT virions were loaded for comparative purposes. Proteins were visualized using antibodies MAbE12-E3 (pUL36), M780 (pUL37), MAb203 (pUL17), and MAb166 (pUL25). The major capsid protein pUL19 was monitored as a loading control using antibody DM165. The positions of molecular weight standards (in thousands) are indicated to the right of each panel.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4300765&req=5

Figure 2: Protein compositions of HSV-1 WT and mutant virus capsids as determined by Western blot analysis of cytoplasmic C-capsids purified from the cytoplasm of cells infected with WT HSV-1 or the inner tegument deletion mutants ARΔUL36 and FRΔUL37. Serial 2-fold dilutions of each sample were run on a 10% polyacrylamide gel and transferred to Hybond ECL nitrocellulose membranes for analysis. Equal amounts of purified WT virions were loaded for comparative purposes. Proteins were visualized using antibodies MAbE12-E3 (pUL36), M780 (pUL37), MAb203 (pUL17), and MAb166 (pUL25). The major capsid protein pUL19 was monitored as a loading control using antibody DM165. The positions of molecular weight standards (in thousands) are indicated to the right of each panel.
Mentions: pUL17 and pUL25, which make up the CVSC, are essential for packaging and retention of DNA in capsids. Therefore, the greatly reduced size of the CVSCs found on ARΔUL36 capsids was surprising, as both the component proteins would be expected to be present on DNA-containing C-capsids. The small size of the CVSC in these cytoplasmic C-capsids suggests either that the archetypal CVSC seen on WT nuclear C-capsids contains, or is stabilized by, pUL36 or that the UL36 mutant capsids contain altered amounts of pUL17 and pUL25. To investigate the latter possibility, we examined the protein composition of the capsids by Western blotting (Fig. 2). Increasing amounts of each capsid preparation were probed with antibodies against pUL17, pUL19, pUL25, pUL36, and pUL37. As expected, pUL36 and pUL37 were not detected in the ARΔUL36 and FRΔUL37 samples, respectively. ARΔUL36 capsids also lacked pUL37, thereby confirming that complex formation with pUL36 is required for addition of pUL37 to capsids (55). pUL36 and pUL37 were both present in WT capsids, but the amounts were appreciably lower than in WT virions. Importantly, the amounts of pUL17 and pUL25 in ARΔUL36 capsids were similar to those for the WT and FRΔUL37, thereby demonstrating that the reduced size of the CVSC was not caused by loss of either of these proteins.

Bottom Line: In addition, the presence of full-length pUL36 results in weak density that extends the CVSC toward the penton, suggesting either that this extra density is formed directly by pUL36 or that pUL36 stabilizes other components of the vertex-tegument interface.Herpesviruses have complex particles that are formed as a result of a carefully controlled sequence of assembly steps.We show that the largest viral protein, pUL36, which occupies the layer of tegument closest to the capsid, is essential for formation of structurally normal connections to the capsid.

View Article: PubMed Central - PubMed

Affiliation: MRC-University of Glasgow Centre for Virus Research, Glasgow, United Kingdom.

Show MeSH
Related in: MedlinePlus